Aerobic bacteria associated with diabetic foot ulcers and their susceptibility pattern

Diabetic foot ulcer presents as open sore in about 15% of diabetics. They are commonly located at the bottom of the foot. Most diabetics’ hospital admission are results of foot ulcer (American Podiatric Medical Association, 2016). Diabetes is a metabolic disorder characterised by hyperglycaemia which results from defects in insulin action, secretion or both. Foot ulcers are complications of uncontrolled diabetes. It could also be due to muscle atrophy, foot deformity, peripheral neuropathy and neuropathic fractures (Raja, 2007).

The prevalence of diabetic foot ulcer is about 13% in North America, 3% in Europe, 7.2% in Africa, 18.1% in Khartoum, Sudan, 15% in Tanzania, 13% in Cameroon and 9.5% in Nigeria (Almobarak et al. 2017).

These result in two thirds of all non-traumatic amputations. Disease presentations include Charcot joint, ulcerations, fracture or gangrene (Krishman et al. 2008; Spichler et al. 2015).

The bacteriology of diabetic foot infection is highly complicated and mostly polymicrobial. It involves both aerobes and anaerobes. Many researchers have presented a picture of mixed infection with aerobic and anaerobic bacteria (Chin, 2013). Some of the aerobic bacteria associated with diabetic foot infection include Staphylococcus aureus, S. saprophyticus, S. epididermis, Streptococcus pyogenes, S. mutans, Pseudomonas aeruginosa, Bacillus subtilis, Proteus species, Escherichia coli and Klebsiella pneumoniae. The anaerobic bacteria include Peptostreptococcus species, Anaerobic Streptococci, Bacteriodes fragilis and Clostridium species (Lipsky et al. 2012; Richard et al. 2012).

Diagnosis is based on combination of signs such as erythema around lesion, local tenderness, local warmth, pus, swelling and indurations. The risk factors are middle or old age, diabetic neuropathy, infection, cigarette smoking, poor glycaemic control, previous foot ulcerations, amputations and ischemia of small and large blood vessels (Scott, 2013; Wu et al. 2007).

Diabetic foot infection can be either superficial or deep. The deep infection involves the muscles, bones, superficial fascia and joints. This includes cellulitis, necrotizing cellulitis and wet gangrene. Once the foot ulcer gets to this stage, the patient is advised on the option of amputation.

The severity of a diabetic foot infection is grouped into four grades depending on their symptoms (Raja, 2007). There are no signs and symptoms in grade 1 wounds. Grade 2 ulcers have lesions on the skin without systemic involvement nor the deep tissues but with any two of the following: erythema > 0.5–2 cm around the ulcer, local indurations, swelling, tenderness, warmth, pain and purulent discharge. Grade 3 ulcers have erythema > 2.0 cm with infection involving structures deeper than the skin and subcutaneous tissues such as osteomyelitis, septic arthritis, or deep abscess. Grade 4 ulcers present with systemic inflammatory respond with at least two of the following signs: temperature > 38 °C, pulse > 90 bpm, PaCO₂ < 32 mmHg, leucocytes > 12,000 or < 4000 per mm³ and 10% of immature (band forms) leucocytes (Raja, 2007).

Pal and Gupta (2016) in their study in Kolkata, India, reported that grade 4 foot ulcers on diabetic patients were commonly infected with mixed variety of bacteria including ESBL Klebsiella species, Methicillin-resistant Staphylococcus aureus (MRSA) and so on. They also reported that patients with very long hospital stay had negative culture reports.

In cases where the foot ulcers have developed, infection may be avoided by maintaining good hygiene in and around the foot ulcer and covering the purulent lesions with a waterproof dressing (Singh et al. 2015). Diabetic foot infections are usually inadequately managed due to poor knowledge on the microbial agents associated with the ulcers. There is a need to investigate the pathogens infecting these ulcers and their susceptibility pattern which may improve the patient’s management and reduce the frequency of amputation. This study was carried out to characterise aerobic bacteria associated with the foot ulcers, determine antimicrobial susceptibility pattern of isolates to commonly used antibiotics and assess the severity of the diabetic foot among subjects in the locality.

Figure 2 shows the distribution of bacterial isolates among subjects. Staphylococcus aureus had the highest degree of occurrence (32 (32.9%)) followed by Pseudomonas aeruginosa (24 (24.7%)).

A 100% infection rate was recorded in the study with a total of 97 isolates. Mono-microbial infection occurred among 14 (28.0%) of the subjects while polymicrobial infection occurred among 36 (72.0%) of the subjects (Fig. 3).

The gram-positive isolates showed higher susceptibility to erythromycin (67.7%), followed by amoxicillin (61.2%) (Table 4). The gram-negative isolates were more susceptible to ciprofloxacin (80.4–100%), followed by zinaceff (25.0–66.7%) and amoxicillin (50.0–64.7%). They exhibited a high resistance to ampicillin (76.5–84.6%), cotrimoxazole (41.2–84.6%) and gentamycin (58.3–75.0%).

In this study, subjects aged 50–59 years were more susceptible to diabetic foot infection. This is in agreement with the report of Karmaker et al. (2016) in Dhaka, Bangladesh with most of their subjects with average age of 58 years and older than 40 years. The mean age of the subjects in this study was 55 ± 10.2 years. The study revealed that diabetics in their fourth and fifth decade of life were more prone to DFU.

A total of 97 aerobic bacterial isolates were encountered in the 50 subjects accounting for an average of 1.94 isolates per subject. This pathogen isolation rate per lesion is lower than the 2.51 reported by Viswanathan et al. (2012) and 2.37 reported by Carvalho et al. (2014) respectively. The lower isolation rate in this study may be due to the incubation method used. Viswanathan et al. (2012) and Carvalho et al. (2014) used both aerobic and anaerobic methods, but only aerobic incubation method was carried out in this study due to lack of equipment and materials.

Staphylococcus aureus was the only gram-positive bacterial isolate 31 (31.9%) while Pseudomonas aeruginosa 24 (24.7%) was the most common gram-negative isolate. Ramakant et al. (2011) in Lucknow, India, also reported P. aeruginosa as the most common gram-negative isolate in diabetic foot ulcer patients in their study.

The polymicrobial infection rate in this study was 36 (72.0%) while the monomicrobial rate was 14 (28.0%). Our findings is higher than the 67.2% reported by Shanmugam et al. (2013) but lower than the 80% and 87.2% polymicrobial infection rates reported by Wright et al. (2011) and Altrichter et al. (2015) respectively. The high polymicrobial infection rate reported in this study point to the fact that most of our subjects suffered severe diabetic foot infections with grade 4 wounds.

Our study reveals an indirect relationship between the duration of diabetic foot ulcer (DFU) and bacterial infections even though there was no statistically significant relationship between DFU and bacterial infections (χ² = 21.1, p ≥ 0.05). Subjects with duration of ulcer ≤ 1 year 29 (58.0%) had the highest burden of bacterial infection 56 (57.7%) while subjects with DFU ≥ 10 years had the lowest bacterial infections. This study did not investigate the reason for the skew in the number of subjects with DFU as the duration of infection increased. The reduction may point to the fact that diabetic subjects with foot ulcers ≥ 10 years may not survive complications of the diabetes which include amputation in our locality.

Antibiotic susceptibility of isolates to commonly used antibiotics was low. None of the isolates showed 100% susceptibility to any of the antibiotic tested. The multiple antibiotic resistance of isolates to quinolones and cephalosporins which are commonly used in our locality calls for an immediate action on the controlled use of antimicrobials in the hospitals and the need to monitor resistance. Good antimicrobial use is necessary for effective wound management. The low susceptibility of isolates observed in this study agrees with the reports of Anguzu and Olila (2007) in Uganda and Ogba et al. (2017) in Nigeria.

A low level of susceptibility of isolates tested with ampicillin and cotrimoxazole was observed. This may be attributed to the fact that ampicillin and septrin have been widely abused and frequently implicated in self-medication in Nigeria. Ogbera et al. (2006) also reported that high level of antibiotic abuse in Nigeria arise from self-medication which is associated with inadequate dosage and failure to comply with treatment regimen. These antibiotics are being sold over the counter with or without prescription.

The study recorded 100% infection rate among the diabetic foot ulcer subjects. All the subjects had grade 4 wounds. The infections were both monomicrobial and polymicrobial. This study showed a dominance of gram-negative bacteria among the isolates. The multiple antibiotic resistance of the bacterial isolates calls for the need to monitor resistance. Best practice is to perform antibiotic susceptibility testing before treatment.